Tests for serum concentration of TNF inhibitors

Test result

Result is given as concentration of TNF inhibitor. Limit of detection: 0.65 Âµg/mL.

Method

The assay is performed with an in vitro reporter cell line (iLite) which measures the amount of drug in the serum that can inhibit TNF-alpha binding to the TNF receptor on the cells (Lallemand 2011 and see figure). The specific TNF-alpha binding activates a reporter gene (Firefly Luciferase) and the effect of the produced enzyme can be measured.

The method is very stable and reproducible and is based on an arrest of the normal cell cycle and the cells can be used straight from the freezer. The cellular activity assay has a very good correlation to classical concentration measurement by radioimmunoassay (RIA) but measures in contrary to binding assays (RIA, ELISA, HMSA etc.) the activity of the drug, which is of a high therapeutic significance and clinical value.

Result interpretation

At detectable levels of the drug and when the patient has an effect of treatment with TNF inhibitors the presence of NAbs are considered to be of low therapeutic significance.

Normal or high concentration of active drug but no treatment effect is observed in patients with a profile of the disease that is not driven by TNF levels.

When secondary failure is suspected despite the presence of normal or high concentrations of active drug, the sample may have been collected not at Cmin. Moreover, NAbs can be measured with precise results only at low drug concentration. If interested to test for NAbs the patient should be taken out of treatment and sampling should be repeated when the drug concentration is considered to be less than 0.65 Âµg/mL.

When the concentration of the TNF inhibitors is below the detection level, this may be caused by NAbs against the TNF blocker which prevents the drug to bind to TNF-alfa. This in turn causes the TNF-alpha to bind normally to the TNF receptor. Determination of NAbs is done in a separate analysis.

Patient compliance problems may be the reason when the serum concentration of TNF inhibitors is lower than expected in combination with undetectable levels of NAbs. Low drug concentration and no treatment effect may also suggest that the patient needs a higher drug dose to achieve the desired therapeutic effect.

Low concentrations of the drug can also be due to non-neutralizing ADA. Non-neutralizing ADA can both limit the bioavailability (with local administration of the drug) and increase the elimination of the drug in the circulation.

The result interpretation is summarised in algorithms that are found here.